Project Summary/Abstract: The first three years-of-life are critically important for establishing growth trajectories and gut microbiota composition, both of which are influenced by diet and other environmental factors. Obesity rates are increasing worldwide and infants who are formula-fed (FF) for the first 6 months of life are ~2.5-times more likely to be obese at 2-years than breast-fed infants (BF); however, little data exists on infants who are both BF and FF (CF). Gut microbial composition is a determinant of obesity risk and microbiota differences exist between BF- vs. FF-fed infants, but less is known about the microbiota of CF infants. Human milk oligosaccharides (HMO) and prebiotics are fermented to short chain fatty acids (SCFA), which signal to the host through the free fatty acid receptors FFAR2 and FFAR3 to influence immune and metabolic function and obesity risk. We have shown that the microbiota and SCFA composition differ between FF and BF infants and that systems biology approaches that combine SCFA and FFAR-linked genes expression in exfoliated intestinal epithelial cells discriminate BF from FF infants. However, it is unknown whether these factors directly influence infant weight gain. The goal of this proposal is to determine how differences in dietary prebiotics influence mutualistic host-microbe interactions in a longitudinal, prospective birth cohort of 440 children and to relate those to infant growth trajectory and weight and body composition at age 3. Our central hypothesis is that dietary HMO and prebiotics produce different microbiome and SCFA composition in HM, FF and CF infants and that SCFA will mediate infant growth trajectories and body composition through interactions with gut FFAR. The proposed experiments will use systems biology approaches to illuminate transgenomic cross- talk between host exfoliated intestinal epithelial cells and the gut microbiota will provide mechanistic insight into the molecular pathways underlying host-microbe interactions in the gut that are associated with infant weight gain and body composition. Two specific aims will be undertaken to test our central hypothesis: 1) Determine the impact of early nutrition on microbiota composition and short chain fatty acid composition and relate those findings to growth trajectories in the first 3 years-of-life and BMI and body composition at age 3.; and 2) Annotate host exfoliated epithelial cell transcriptome and bacterial metatranscriptome profiles and elucidate host/commensal relationships focusing on FFAR-linked pathways and relate those findings to growth trajectories in the first 3 years-of-life and BMI and body composition at age 3. Our team is ideally positioned to undertake this research. Our pioneering noninvasive approach that simultaneously monitors gene expression in exfoliated epithelial cells and gut microbiota of infants is highly innovative. This body of work is significant because molecular biomarkers that define the relationship between dietary intake, microbiota composition, host gene expression and child health outcomes will be identified. These host-microbial molecular finger prints will enable patient-powered precision medicine to optimize infant growth and reduce childhood obesity risk.